3,8-Dioxo-scirpen-4β,15-diol esters and their use as antitumor agents

ABSTRACT

A novel series of 3,8-dioxo-scirpen-4 beta ,15-diol esters is provided for use as antitumor agents. Also provided are various novel intermediates used in the production of such compounds.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to novel trichothecene derivatives, toprocesses for their production and to their use as antitumor agents forthe inhibition of malignant tumors in mammals.

2. Description of the Prior Art

The trichothecene derivatives of the present invention all contain a9,10 double bond and a 12,13-epoxy function. The basic skeleton andnumbering system for this class of trichothecenes is shown below.##STR1## Various examples of both naturally occurring and semisyntheticcompounds of this class have been described in the literature.Illustrative of the more relevant publications are the following:

1. The compound anguidine (also called diacetoxyscirpenol) having theformula ##STR2## is disclosed as an antitumor agent in U.K. Pat. No.1,063,255. Phase I clinical trials of anguidine in the United Stateshave been reported in Proc. Amer. Assoc. Cancer Res. 17:90 (1976) andProc. Amer. Assoc. Cancer Res. 18:296 (1977). Also disclosed (at leastgenerically) are various derivatives of anguidine such as anguidol (alsocalled scirpentriol or 3α,4β,15-trihydroxy-12,13-epoxytrichothec-9-ene),monodesacetylanguidine (presumably15-acetoxy-3α,4β-dihydroxy-12,13-epoxytrichothec-9-ene ormonoacetoxyscirpendiol) and esters of anguidine, anguidol andmonodesacetylanguidine.

Monoacetoxyscirpenol and various esters of scirpentriol are alsodisclosed in J. Agric. Food Chem. 24(1): 97-103 (1976) as mycotoxins.

2. Japanese Published Applications Nos. J4 9,134,891 and J4 9,134,892disclose T2 and HT2 toxins of the formula ##STR3## wherein R is --OH or##STR4## The compounds are said to be useful as antiviral agents.

3. U.S. Pat. No. 4,129,577 discloses anguidine derivatives of theformula ##STR5## wherein R₁ is H or ##STR6## and R is an alkyl oraromatic group or is an acyl group ##STR7## in which R¹ is an aliphatic,cycloaliphatic or aromatic group or a carbamate group --CONH--R¹. Thecompounds are useful as cytotoxic agents.

4. Neosolaniol having the formula ##STR8## wherein the 8-hydroxy groupis of the α-configuration is disclosed in J. Pure and Applied Chemistry35(3):309 (1973) as a mycotoxin and inhibitor of protein synthesis.

5. U.S. Pat. No. 3,428,652 discloses anguidine derivatives of theformula ##STR9## wherein R₁ is H and R₂ is methyl or, R₁ and R₂ togetherrepresent propylene, and Hal is Cl, Br or I. The compounds are reportedto have antitumor activity.

6. Toxins isolated from culture filtrates of F. scirpi and having theformula ##STR10## are disclosed in J. Chem. Soc (C), 375 (1970).

7. Trichothecene derivatives of the formula ##STR11## wherein R¹, R², R³and R⁴ are --OH or --OCOCH₃ are disclosed in Biochemical Pharmacology24:959-962 (1972) as having larvicidal activity. The degree of activityis said to be greatest in the compound where R¹ ═R² ═R³ ═R⁴ ═OH andleast in the fully acetylated compound. It is suggested in thepublication that the order of cytotoxic activity in this series is thesame as the order of larvicidal activity.

8. The 12,13-epoxytrichothecenes of the general formula ##STR12##wherein R¹ and R³ are H, OH or esterified OH and R² is OH, ═O oresterified OH are described in Biochemical and Biophysical ResearchCommunications 57(3): 838-844 (1974) as inhibitors of protein synthesis.The publication indicates that substitution of a carbonyl group at theC-8 position of the above compounds results in a moderate loss ofactivity and that substitution of a carbonyl group at R² results incomplete loss of activity.

9. Helvetica Chimica Acta 48:962-988 (1965) discloses scirpentriol,various acetoxy esters of scirpentriol and the compound of the formula##STR13## No biological properties of the 3-keto compound are indicated.

SUMMARY OF THE INVENTION

The present invention provides novel trichothecene derivatives of thegeneral formula ##STR14## wherein R¹ and R² are each independently(lower)alkyl; halo(lower)alkyl; alkenyl of the formula --CR³ ═C⁴ R⁵ inwhich R³ is hydrogen, (lower)alkyl or 1'-halo(lower)alkyl and R⁴ and R⁵are each independently hydrogen or (lower)alkyl; alkynyl of the formula--C.tbd.CR⁶ in which R⁶ is hydrogen or (lower)alkyl; or a radical of theformula

    Ar--(CH.sub.2).sub.m --

in which m is 0 or an integer from one to four and Ar is ##STR15##wherein R⁷, R⁸ and R⁹ are each independently hydrogen, halogen,(lower)alkyl or (lower)alkoxy.

In another aspect the present invention provides compounds of thegeneral formula ##STR16## wherein R¹ and R² are as disclosed for thecompounds of formula I, with the provisos that (1) R¹ and R² in formulaII may not both be methyl and (2) when R¹ and R² in formula III ismethyl, the 8-hydroxy group must be in the β-configuration.

The compounds of formulae I-III are antitumor agents for treatment ofmalignant tumors in mammals. Compounds of formulae II and III are alsointermediates useful in the preparation of the compounds of formula I.

DETAILED DESCRIPTION

The various substituent groups disclosed above in connection with thenovel compounds of the present invention may be further defined asfollows:

(a) Halo or halogen includes chlorine, bromine, fluorine and iodine;

(b) (Lower)alkyl includes both straight and branched chain saturatedaliphatic hydrocarbon radicals having from 1 to 4 carbon atomsinclusive, e.g. methyl, ethyl, n-propyl, isopropyl or n-butyl;

(c) (Lower)alkoxy includes C₁ -C₄ alkoxy radicals, the alkyl portion ofsuch radicals being defined as in (b) above. Examples include methoxy,ethoxy, n-propoxy, isopropoxy and n-butoxy;

(d) Halo(lower)alkyl includes (lower)alkyl radicals as defined under (b)where one or more hydrogen atoms are substituted by halogen as definedunder (a). Examples include --CF₃, --CCl₃, --CH₂ Cl, --CHCl₂, --CH₂ CH₂Cl, --CH₂ CF₃, --CH₂ CH₂ CHClCH₃ or --CH₂ CHClCH₂ CH₃ ; and

(e) The phenyl and naphthyl groups above may be optionally substitutedby one, two or three nonhydrogen substituents at any of the availablepositions of the ring system. The naphthyl radical may be either the α-or β-isomer. Preferred aryl radicals are those which are unsubstitutedor which have one non-hydrogen substituent.

As denoted by the structural formulae provided above, the compounds offormulae II and III all have the 3-hydroxy substituent in theα-configuration. Additionally, the compounds of formulae I-III all havethe β-configuration at the 4-substituent. Compounds of formula III mayexist in the form of the 8α- and 8β- hydroxy epimers or mixturesthereof, but the present invention is specifically limited to the 8βepimer of the compound where R¹ ═R² ═methyl. Certain compounds withinthe scope of formulae I-III contain asymmetric carbon atoms (e.g. whenR¹ and R² are butyl) and, in such cases, the compounds may exist in theform of the individual optical isomers as well as the racemates.

The compounds of formula I may be prepared by reacting the appropriatediol intermediate of formula III with about two equivalents of asuitable oxidizing agent in an inert organic solvent.

In general any oxidizing agent capable of converting a stericallyhindered hydroxyl group to a carbonyl group may be employed in the aboveprocess. A particularly preferred reagent is dimethylsulfoxide-trifluoroacetic anhydride (DMSO-TFAA) which is described in J.Org. Chem. 41(20): 3329 (1976). This reagent may be conveniently used ina dry inert organic solvent such as methylene chloride ortetrahydrofuran at temperatures of from about -78° C. to -50° C. Uponaddition of the reagent to the diol intermediate III, adimethylalkoxysulfonium salt is formed which on treatment with base(e.g. triethylamine) is rapidly converted in good yield to thecorresponding diketo product I. An alternative preferred oxidizingreagent is dimethylsulfoxide-acetic anhydride. This oxidizing agent isemployed in a similar manner as DMSO-TFAA except that the reaction ispreferably carried out at about room temperature.

Compounds of formula I may also be prepared from the appropriate 8-ketointermediate of formula II by oxidation with about one equivalent of asuitable oxidizing agent such as dimethyl sulfoxide-trifluoroaceticanhydride, dimethylsulfoxide-acetic anhydride or N-chlorosuccinimidedimethylsulfide in an inert organic solvent such as methylene chlorideor toluene. The preferred temperature conditions for this reaction areabout -78° to -50° C. for dimethyl sulfoxidetrifluoroacetic anhydride,room temperature for dimethylsulfoxide-acetic anhydride and 0° C. forN-chlorosuccinimide dimethyl sulfide.

Other reaction temperatures may be successfully employed in theoxidations of compounds II and III, but product yields may be reducedfrom those achieved under the preferred conditions.

The 8-keto intermediate of formula II may be prepared by selectiveoxidation of the 8-hydroxy group of the appropriate diol compound offormula III in an inert organic solvent such as dioxane, acetone ormethylene chloride. Mild oxidizing agents are used in this conversionsuch as manganese dioxide, 2,3-dichloro-5,6-dicyano-1,4-benzoquinone orpyridinium chlorochromate. The most preferred oxidizing agent for thisstep is pyridinium chlorochromate. Advantageous results are obtainedwith this reagent when the oxidation reaction is carried out at aboutroom temperature in an acetate-buffered methylene chloride solvent (seeTetrahedron Letters 31:2647-2650 (1975) for further details of theprocedure).

Diol intermediates of formula III may be prepared from the appropriate3α-hydroxy ester of the formula ##STR17## by selectively oxidizing esterIV in an inert solvent so as to introduce an 8-hydroxy group. Apreferred oxidizing agent is selenium dioxide which may be convenientlyreacted with ester IV in aqueous dioxane at reflux temperature.

Alternatively, the 8-hydroxy group of intermediate III may be introducedon the appropriate ester of formula IV by subjecting ester IV toN-bromosuccinimide bromination in an inert organic solvent such asmethylene chloride and then displacing the 8-bromo group with aqueousbase in the presence of a silver salt such as silver trifluoroacetate.

Starting material 3α-hydroxy esters of general formula IV are known inthe art or are prepared by methods well-known to those skilled in theart. Examples of suitable methods are provided below under "Preparationof Starting Materials", but in general the esters may be prepared asshown in the following schemes: ##STR18##

EXPLANATION OF SCHEMES I-III

Using anguidine as the starting material, other 4β,15-diacylated estersof formula IV may be prepared by protecting the 3-OH group as byconversion to a tetrahydropyranyl ether (1), and then subjecting the3α-THP derivative to partial basic hydrolysis to give a mixture of the4-OH (3) and 4,15-OH (2) derivatives.

Compound 2 may then be acylated in accordance with conventional methodswith about two equivalents of a suitable acylating derivative of acarboxylic acid R--COOH to produce a 4,15-diacylated derivative 4 whichmay then be de-protected to give 5. The acylation is typically carriedout with an acid halide or acid anhydride, preferably in the presence ofan organic base such as pyridine or lutidine. Scheme II results information of a 4,15-diacylated ester of general formula IV having R¹=R².

To prepare esters of formula IV where R¹ ≠R², the 4,15-diol 2 may beacylated with less than two equivalents of acylating agent to give amixture of monoacylated derivatives 6 and 7 as shown in Scheme III.These derivatives can be separated chromatographically and then treatedwith a second acylating agent to give the diacylated derivatives 8 and8'. Upon de-protection the products 9 and 9' containing mixed acylgroups are produced.

Mixed diacylated esters of formula IV where R¹ is methyl may also beprepared by acylation and de-protection of compound 3.

BIOLOGICAL ACTIVITY

Representative compounds of the present invention were tested forantitumor activity against the transplantable mouse tumors P-388leukemia, L-1210 leukemia and Lewis Lung carcinoma and the results ofthese tests are shown below in Tables I-VI. The methodology usedgenerally followed the protocols of the National Cancer Institute [see,for example, Cancer Chemotherapy Rep., Part 3, 3:1-103 (1972)]. Theessential experimental details are give at the bottom of the tables.

                  Table I                                                         ______________________________________                                        Effect of Compound of Example 1 on P-388 Leukemia                                                      Effect                                                                              Average                                                  Dose    MST    MST   Weight Survivors                               Material  mg/kg   Days   % T/C Change,g                                                                             Day 5                                   ______________________________________                                        Anguidine 1.6     15.5   172   +1.0   6/6                                               0.8     14.5   161   +0.6   6/6                                               0.4     13.0   144   -0.6   6/6                                               0.2     12.0   133   -1.3   6/6                                               0.1     10.0   117   +0.5   6/6                                               0.05    10.0   111   +0.6   6/6                                     Compound                                                                      of Example 1                                                                            6.4     Tox    Tox   Tox    0/6                                               3.2     Tox    Tox   Tox    1/6                                               1.6     Tox    Tox   Tox    1.6                                               0.8     15.0   167   -0.8   5/6                                               0.4     14.0   156   +0.3   6/6                                               0.2     12.0   133   +0.7   6/6                                               0.1     12.0   133   +0.7   6/6                                               0.05    10.0   111   +0.9   6/6                                     Control   Saline   9.0   --    +2.4   10/10                                   ______________________________________                                         Tumor inoculum: 10.sup.6 ascites cells implanted i.p.                         Host: CDF.sub.1 ♀ mice.                                                Treatment: Once daily for 9 days starting day 1                               Evaluation: MST = median survival time.                                       Effect: % T/C = (MST treated/MST control) × 100.                        Criteria: T/C ≧ 125 considered significant antitumor activity.    

                  Table II                                                        ______________________________________                                        Effect of Compound of Example 3 on L-1210 Leukemia                                                      Effect                                                                              Average                                                Dose, IP  MST    MST   Weight Survivors                              Material mg/kg/day Days   % T/C Change,g                                                                             Day 5 (30)                             ______________________________________                                        Anguidine                                                                              2.0       11.0   157   -0.8   6/6                                    NSC 141537                                                                             1.6       11.0   157   -0.3   6/6 (1/6)                                       1.2       11.0   157   -0.3   6/6                                             0.8       11.0   157   -0.3   6/6                                             0.4       10.0   143   -0.1   6/6                                             0.2       9.0    129   +0.5   6/6 (1/6)                              Compound of                                                                   Example 3                                                                              2.0       9.0    129   -1.6   3/6                                             1.6       16.5   236   -1.3   6/6                                             1.2       13.5   193   -1.9   6/6                                             0.8       12.0   171   -1.0   6/6                                             0.4       11.5   164   -0.9   6/6 (1/6)                                       0.2       11.0   157   -0.5   6/6                                             0.1       8.0    114   -0.6   6/6 (1/6)                              Control  Saline    7.0    --    +0.9   10/10                                  ______________________________________                                         Tumor inoculum : 10.sup.6 ascites cells implatned ip                          Host : BDF.sub.1 ♀ mice.                                               Treatment : Once daily for 9 days starting day 1                              Tox : <4/6 mice alive on Day 5                                                Evaluation : MST = median survival time.                                      Effect : % T/C = (MST treated/MST control) × 100.                       Criteria : % T/C ≧ 125 considered significant antitumor activity. 

                  Table III                                                       ______________________________________                                        Effect of Compound of Example 3 on P-388 Leukemia                                                       Effect                                                                              Average                                                Dose      MST    MST   Weight Survivors                              Material mg/kg/day Days   % T/C Change,g                                                                             Day 5                                  ______________________________________                                        Anguidine                                                                              1.6       16.0   178   +1.3   6/6                                    NSC 141537                                                                             0.8       15.0   167   -0.5   6/6                                             0.4       13.0   144   +0.6   6/6                                             0.2       12.0   133   +0.2   6/6                                             0.1       12.0   133   +0.9   6/6                                             0.05      11.0   122   +0.9   6/6                                    Compound of                                                                   Example 3                                                                              6.4       8.0     89   -0.7   3/6                                             3.2       14.5   161   -2.5   4/6                                             1.6       22.5   250   -0.9   6/6                                             0.8       17.0   189   +0.2   6/6                                             0.4       14.5   161   -0.2   6/6                                             0.2       13.5   150   -1.0   6/6                                             0.1       12.5   139   -0.3   6/6                                             0.05      12.0   133   -1.5   6/6                                    Control  Saline    9.0    --    +0.8   10/10                                  ______________________________________                                         Tumor inoculum : 10.sup.6 ascites cells implanted i.p.                        Host : CDF.sub.1 ♂ mice.                                                 Treatment : Once daily for 9 days starting day 1                              Tox : <4/6 survivors Day 5.                                                   Evaulation : MST = median survival time                                       Effect : % T/C = (MST treated/MST control) × 100.                       Criteria : % T/C ≧ 125 considered significant antitumor activity. 

                  Table IV                                                        ______________________________________                                        Effect of Compound of Example 3 on P-388 Leukemia                             (repeat test)                                                                                                            Sur-                                               Dose          Effect                                                                              Average                                                                              vivors                                     Treat-  mg/kg/   MST  MST   Weight Day 5                              Material                                                                              ment    inj      Days % T/C Change,G                                                                             (30)                               ______________________________________                                        Anguidine                                                                             Days    2.0      16.0 178   +0.5   6/6                                        1→9                                                                            1.6      16.5 183   +0.3   6/6                                                1.2      15.5 172   +0.9   6/6                                                0.8      14.0 156   +0.7   6/6                                Compound                                                                      of                                                                            Example 3                                                                             Days    2.0      10.0 111   -0.3   6/6                                        1→9                                                                            1.6      27.5 306   +0.1   6/6 (2)                                            1.2      19.5 217   +0.8   6/6                                                0.8      19.5 217   -0.1   6/6                                        Day 1   10.0     13.5 150   -1.8   4/6                                        only    7.0      12.5 139   -0.6   4/4                                Control         Saline   9.0  --    +1.0   10/10                              ______________________________________                                         Tumor inoculum : 10.sup.6 ascites cells implanted i.p.                        Host : CDF.sub.1 ♂ mice.                                                 Treatment : Once daily for 9 days starting day 1                              Tox : < 4/6 survivors Day 5.                                                  Evaulation : MST = median survival time                                       Effect : % T/C = (MST treated/MST control) × 100.                       Criteria : % T/C ≧ 125 considered significant antitumor activity. 

                  Table V                                                         ______________________________________                                        Effect of Compound 5 on P-388 Leukemia                                                                  Effect                                                                              Average                                                Dosage, IP                                                                              MST    MST   Weight Survivors                              Material mg/kg/inj Days   % T/C Change,g                                                                             Day 5(30)                              ______________________________________                                        Anguidine                                                                              1.6       16.0   200   +0.9   6/6                                    NSC 141537                                                                             0.8       14.0   175   +1.6   6/6                                             0.4       13.0   163   +1.6   6/6                                             0.2       12.0   150   +0.9   6/6                                    Compound of                                                                   Example 5                                                                              6.4       TOX    TOX   TOX    2/6                                             3.2        8.0   100   -0.9   5/6                                             1.6       21.5   270   0      6/6 (1)                                         0.8       18.5   231   +0.3   6/6                                             0.4       15.5   194   +0.6   6/6                                             0.2       12.0   150   +0.4   6/6                                             0.1       12.5   156   +0.5   6./6                                            0.05      12.0   150   +0.1   5/6                                    Control  Saline     8.0   --    +1.8   10/10                                  ______________________________________                                         Tumor inoculum : 10.sup.6 ascites brei cells implanted ip.                    Host : CDF.sub.1 ♀ mice.                                               Treatment : Once daily for 9 days starting day 1                              Tox : <4/6 mice alive on Day 5.                                               Evaluation : MST = median survival time.                                      Effect : % T/C = (MST treated/MST control) × 100.                       Criteria : % T/C ≧ 125 considered significant antitumor activity. 

                  Table VI                                                        ______________________________________                                        Effect of Trichothecene Derivatives                                           on Lewis Lung Carcinoma                                                                Dose, IP         Effect                                                                              Average                                                mg/kg/   MST     MST   Weight Survivors                              Material day      Days    % T/C Change Day 5(60)                              ______________________________________                                        Anguidine                                                                              1.6      21.0    124   +2.2   10/10                                  NSC 141537                                                                             0.8      21.0    124   +1.8   10/10                                           0.4      23.0    135   +1.4   10/10                                  Compound of                                                                            2.0      30.0    176   +0.3   10/10                                  Example 5                                                                              1.5      31.0    182   +0.8   10/10                                           1.0      >60.0   >353  +1.0   10/10 (5)                                       0.5      >60.0   >353  +1.6   10/10 (5)                              Control  Saline   17.0    --    -0.6   10/10                                  ______________________________________                                         Tumor inoculum : 10.sup.6 tumor brei cells, ip.                               Host : BDF.sub.1 ♂ mice.                                                 Treatment : QD 1→9.                                                    Tox : <6/10 mice alive on Day 5.                                              Evaluation : MST = median survival time.                                      Effect : % T/C = (MST treated/MST control) × 100.                       Criteria : % T/C ≧ 125 considered significant antitumor activity. 

Analysis: The compound of Example 1 has an optimum dose (O.D.) of 0.8mg/kg/day vs. 1.6 mg/kg/day for anguidine. This compound thus has twicethe potency of anguidine.

The compound of Example 3 has approximately the same potency asanguidine but is clearly superior in causing prolonged survival of micebearing L-1210 leukemia and in repeated tests with P-388 leukemia.

The compound of Example 5 is about equal in potency to anguidine andcauses greater survival increases in tests on P-388 leukemia and LewisLung carcinoma.

The experimental animal tests described above demonstrate that thecompounds of the present invention possess marked inhibitory actionagainst mammalian malignant tumors. While all of the compounds includedwithin the scope of formulae I-III possess antitumor activity, the mostpreferred compounds for therapeutic use are the compounds of formula I.Compounds of formulae II and III, therefore, are of most importance asintermediates for preparation of the preferred compounds I.

According to another aspect of this invention, therefore, there isprovided a method for therapeutically treating a mammalian host affectedby a malignant tumor which comprises administering to said host aneffective tumor-inhibiting dose of a compound of formula I, II or III.

In yet another aspect of the invention, a pharmaceutical composition isprovided which comprises an effective tumor-inhibiting amount of acompound of formulae I-III in combination with an inert pharmaceuticallyacceptable carrier or diluent. These compositions may be made up in anypharmaceutical form appropriate for parenteral administration.

Preparations according to the invention for parenteral administrationinclude sterile aqueous or nonaqueous solutions, suspensions oremulsions. They may also be manufactured in the form of sterile solidcompositions which can be dissolved in sterile water, physiologicalsaline or some other sterile injectable medium immediately before use.

It will be appreciated that the actual preferred dosages of thecompounds of the present invention will vary according to the particularcompound being used, the particular composition formulated, the mode ofadministration and the particular situs, host and disease being treated.Many factors that modify the action of the drug will be taken intoaccount by those skilled in the art, e.g. age, body weight, sex, diet,time of administration, route of administration, rate of excretion,condition of the host, drug combinations, reaction sensitivities andseverity of the disease. Optimal dosages for a given set of conditionscan be ascertained by those skilled in the art using conventional dosagedetermination tests in view of the experimental animal data provided,the available data on clinical use of anguidine and the above-mentionedguidelines.

The following examples are not limiting but are intended to beillustrative of this invention. SKELLYSOLVE B is a commerciallyavailable petroleum solvent (Skelly Oil Co.) comprising isomeric hexanesand having a boiling point of 60°-68° C. The main component ofSkellysolve B is n-hexane. Unless otherwise indicated, all temperaturesbelow are in degrees Celsius and all solvent percentages are by volume.

PREPARATION OF STARTING MATERIALS Preparation 14β,15-Diacetoxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene

A mixture of 4β,15-diacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene(12.81 g, 35 mmol), 2,3-dihydro-4H-pyran (17.5 ml, 189 mmol), andp-toluenesulfonic acid (70 mg, 0.35 mmol) in 150 ml of CH₂ Cl₂ wasstirred at room temperature for 2 h. After addition of 2.1 g of K₂ CO₃,the reaction mixture was diluted with 400 ml of CH₂ Cl₂ and washed withsaturated NaHCO₃ solution and brine. Drying over K₂ CO₃ and removal ofthe solvent gave a colorless oil which crystallized slowly frompetroleum ether to give 11.30 g (72%) of solid. m.p. 93°-94° C.;IR(KBr): 2976, 1746, 1249, 1080, 1040, 988 cm⁻¹.

Anal. Calc'd for C₂₄ H₃₄ O₈ : C, 63.98; H, 7.61. Found: C, 64.35; H,7.58.

Preparation 2 3α,4β,15-trihydroxy-12,13-epoxytrichothec-9-ene

4β,15-diacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene (15 g) wasstirred for 20 minutes in 300 ml of methanol with 900 ml of 0.3 N sodiumhydroxide (Sigg et al. Helv. Chim. Acta, 48, 962-988 (1965). Thesolution was placed on a column containing 1 kg of DOWEX 50 (H⁺ cycle)prepared with 20% methanol in water. The column was eluted with 3 l ofthe same solvent, the eluate concentrated, and the residual aqueoussolution freeze-dried. The powder was dissolved in methanol, mixed with10 g of silica gel, and dried in vacuo. The dry silica gel mixture wasplaced on a column of fresh silica gel (2.5×100 cm) and eluted withmethylene chloride with increasing amounts of methanol. Fractionsappearing homogeneous on TLC plates were dried and crystallized fromethyl acetate. Yield: 7.3 g, m.p. 194°-195° C. IR(KBr): 3490, 3450,3390, 2990-2900 (four peaks), 1675, 960 and 950 cm⁻¹. [α]_(D) ²² =-15.4°(c=1, acetone).

Anal. Calc'd for C₁₅ H₂₂ O₆ : C, 63.81; H, 7.86. Found: C, 63.71; H,7.80.

Alternatively, the 3-O-tetrahydropyranyl derivative (Preparation 3below) (1 g) was stirred for four hours in 115 ml of 95% ethanol and 23ml of 1 N HCl. The solution was azeotropically distilled with theaddition of absolute ethanol, the concentrated ethanolic solutiondiluted with diethyl ether, and the resulting title product crystallizedfrom ethyl acetate as a gum.

Preparation 34β,15-Dihydroxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene

To a solution of4β,15-diacetoxy-3-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene(1.067 g, 2.37 mmol) in 40 ml of tetrahydrofuran and methanol (5:3 v/v)was added 20 ml of 0.3 N NaOH solution. After 2.5 h of stirring at roomtemperature, an additional 20 ml of 0.3 N NaOH solution was introduced,and stirring was continued for 18.5 h. The resulting solution wasdiluted with CH₂ Cl₂ (200 ml) and washed with water. The aqueous layerwas reextracted with CH₂ Cl₂ (2×50 ml). The combined CH₂ Cl₂ layers werewashed with brine and dried over K₂ CO₃. Removal of the solvent gave 891mg of foam, which was subsequently chromatographed on silica gel.Elution with 1% methanol-CH₂ Cl₂ gave 46 mg (5%) of15-acetoxy-4β-hydroxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene.A further elution with 5% methanol-CH₂ Cl₂ gave 808 mg (93%) of thetitle compound as an amorphous solid. IR(KBr): 3457, 2943, 1445, 1135,1125, 1078, 1035, 1020, 978, 957 cm⁻¹.

Preparation 415-Acetoxy-4β-hydroxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene

To a solution of4β,15-diacetoxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene(31.14 g, 69.2 mmol) in 800 ml of methanol and tetrahydrofuran (1:1 v/v)was added 400 ml of 1.31 N NH₄ OH solution. After 3 days stirring atroom temperature, 10 ml of concentrated NH₄ OH solution was added to thereaction mixture. Stirring was continued for an additional 4 days. Thevolume of the resulting solution was reduced to 500 ml. Extraction withCH₂ Cl₂, washing with brine, and removal of the solvent gave 37 g of aslightly yellow oil. Chromatography on silica gel (elution with 1%methanol-CH₂ Cl₂) gave 10.65 g (38%) of the title compound as an oil.The NMR and IR spectra of this material were consistent with thestructure of the title compound. IR(KBr): 3430, 2970, 2950, 2875, 1744,1720, 1270, 1248, 1126, 1080, 1040, 972 cm⁻¹.

Preparation 5 4β,15-Bis-(2'-bromoacetoxy)-3α-hydroxy-12,13-epoxytrichothec-9-ene

A solution containing 183 mg (0.5 mmol) of3α-O-(2'-tetrahydropyranyl)-4β,15-dihydroxy-12,13-epoxytrichothec-9-eneand 268 mg (2.5 mmol) of 2,6-lutidine was cooled to 5° C. To thissolution was added with stirring 253 mg (1.25 mmol) of bromoacetylbromide and the resulting solution was kept without cooling for 18 h.The usual work-up gave a gum which was dissolved in 36 ml of 95% ethanoland 6 ml of 1 N HCl and stored for 20 h at 22° C. The usual work-up gavea foam which was chromatographed on 20 g of silica gel (MallinckrodtSILICAR CC-7) using 1% methanol in CH₂ Cl₂ as the solvent. The producteluted as a pale yellow band which gave a foam on evaporation of thesolvent. Crystallization from diethyl ether afforded a colorless solidof m.p. 125°-126°. IR(KBr): 3480, 2960, 1750, 1735, 1280, 1165 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₄ Br₂ O₇ : C, 43.53; H, 4.61. Found: C, 45.47; H,4.84.

Preparation 64β,15-Bis(chloroacetoxy)-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene

A mixture of4β,15-dihydroxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene(808 mg, 2.21 mmol), chloroacetic anhydride (1.132 g, 6.62 mmol) andpyridine (894 mg, 11.05 mmol) in 100 ml of CH₂ Cl₂ was stirred at roomtemperature for 14 h. The reaction mixture was diluted with 200 ml ofCH₂ Cl₂ and washed with saturated NaHCO₃ solution, 1% HCl solution andbrine. Drying over K₂ CO₃ and removal of the solvent gave 1.058 g (92%)of foam which was homogeneous on TLC. A portion of this material waspurified by silica gel chromatography (elution with 0.5% methanol-CH₂Cl₂) to furnish an analytical sample of title product. IR(KBr): 2955,1762, 1740, 1290, 1186, 1172, 1129, 1080, 1039, 977 cm⁻¹.

Preparation 74β,15-Bis(chloroacetoxy)-3α-hydroxy-12,13-epoxytrichothec-9-ene

To a solution of4β,15-bis(chloroacetoxy)-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene(858 mg, 1.65 mmol) in 100 ml of 95% ethanol was added 19 ml of 1 N HClsolution. The resulting solution was stirred at room temperature for 24hours. The reaction mixture was diluted with CH₂ Cl₂ (300 ml) and washedwith saturated NaHCO₃ solution and brine. Drying over K₂ CO₃ -Na₂ SO₄and removal of the solvent gave 600 mg of foam. Chromatography of thismaterial on silica gel (elution with 1% methanol-CH₂ Cl₂) gave 524 mg(73%) of4β,15-bis(chloroacetoxy)-3α-hydroxy-12,13-epoxytrichothec-9-ene. Ananalytical sample was obtained by recrystallization fromchloroform-diethyl ether, m.p. 139°-141° C. IR(KBr): 3450, 2970, 2913,1758, 1742, 1327, 1293, 1192, 1173, 1083, 1008, 967 cm⁻¹. Elution of thesilica gel column with 2% methanol-CH₂ Cl₂ gave 110 mg (15%) of15-chloroacetoxy-3α,4β-dihydroxy-12,13-epoxytrichothec-9-ene.

Anal. Calc'd. for C₁₉ H₂₄ O₇ Cl₂ : C, 52.42; H, 5.56. Found: C, 52.31;H, 5.34.

Preparation 815-Acetoxy-4β-chloroacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene

A mixture of15-acetoxy-4β-hydroxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene(785 mg, 1.92 mmol), chloroacetic anhydride (492 mg, 2.88 mmol) andpyridine (0.309 ml, 3.84 mmol) in 100 ml of CH₂ Cl₂ was stirred at roomtemperature for 28.5 h. The reaction mixture was diluted with CH₂ Cl₂(200 ml) and washed with saturated NaHCO₃ solution and brine. Dryingover K₂ CO₃ and removal of the solvent gave 931 mg (100%) of a whitefoam. The NMR and IR spectra of this material were consistent with thestructure of15-acetoxy-4β-chloroacetoxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene.

To a solution of15-acetoxy-4β-chloroacetoxy-3α-O-(2'-tetrahydropyranyl)-12,13-epoxytrichothec-9-ene(900 mg, 1.98 mmol) in 100 ml of 95% ethanol was added 19 ml of 1 N HClsolution. After 10 h of stirring at room temperature, 200 ml of CH₂ Cl₂was added to the reaction mixture. The aqueous layer which separated wasextracted with 25 ml of CH₂ Cl₂. The combined CH₂ Cl₂ layers were washedwith saturated NaHCO₃ solution and brine. Drying over K₂ CO₃ Na₂ SO₄ andremoval of the solvent gave 662 mg of foam. Chromatography on silica gel(elution with 0.5% methanol-CH₂ Cl₂) gave 350 mg (44%) of the titlecompound. Recrystallization from diethyl ether furnished the analyticalsample; m.p. 166°-167.5° C. IR(KBr): 3500, 3040, 3020, 2989, 2918, 1754,1736, 1378, 1330, 1260, 1208, 1167, 1074, 1052, 960 cm⁻¹ .

Anal. Calc'd for C₁₉ H₂₅ O₇ Cl: C, 56.83; H, 6.29. Found: C, 57.12; H,6.29.

Preparation 9 4β,15-Diacetoxy-3-hydroxy-12,13-epoxytrichothec-9-en-8-one

A mixture of 4β,15-diacetoxy-3α,8β-dihydroxy-12,13-epoxytrichothec-9-ene(270 mg, 0.71 mmol), pyridium chlorochromate (2.23 mg, 1.03 mmol), andanhydrous sodium acetate (29 mg, 0.35 mmol) in 15 ml of CH₂ Cl₂ wasstirred at room temperature for 2.5 h. After filtration through CELITE,the solvent was removed under reduced pressure to give 360 mg of oil.Chromatography on silica gel (elution with 0.75% methanol-CH₂ Cl₂) gave218 mg (81%) of foam which was homogeneous on TLC (thin layerchromatography). IR (KBr): 3445, 2980, 2920, 1740, 1678, 1366, 1240,1042 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₄ O₈ : C, 59.99; H, 6.36. Found: C, 59.36; H,6.41.

Preparation 1015-(trans-2'-butenoyloxy)-3α,4β-dihydroxy-12,13-epoxytrichothec-9-ene

A solution containing 366 mg (1 mmol) of3α-O-(2'-tetrahydropyranyl)-4β,15α-dihydroxy-12,13-epoxytrichothec-9-eneand 395 mg (5 mmol) of dry pyridine in 50 ml of CH₂ Cl₂ (previouslydried over 4 A molecular sieves) was cooled at 5° C. To the stirredsolution was added dropwise 261 mg (2.5 mmol) of trans-2-butenoic acidchloride and, after completion of the addition, the mixture was stirredfor 1 h at 5° C. and for 16 h at ambient temperature. The solution wasdiluted with 50 ml of CH₂ Cl₂ and was successively washed with saturatedaqueous NaHCO₃, brine, 1% aqueous HCl and brine. The organic phase wasdried over Na₂ SO₄ and the solvent was evaporated under reduced pressureto provide 360 mg of a gum. This was dissolved in 50 ml of 95% ethanoland to it was added 5 ml of 2 N HCl. After the solution had been storedat room temperature for 22 h, it was diluted with 100 ml of CH₂ Cl₂ andwas washed successively with H₂ O, saturated aqueous NaHCO₃ and brine.The organic phase was dried over Na₂ SO₄ and the solvent was evaporatedunder reduced pressure to provide 260 mg of gum. This waschromatographed on 20 g of silica gel (Mallinckrodt SILICAR CC-7) using1% methanol in CH₂ Cl₂ as the solvent. The first product eluted was thebis-ester (26 mg) (Preparation 11) followed by 22 mg of the 4-monoester(Preparation 11) and then 147 mg of the title compound which wascrystallized from CHCl₃ -SKELLYSOLVE B as a white solid of m.p. 83°-86°C. IR(KBr): 3440, 2970, 1725, 1190, 1110, 1085, 965 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₆ O₆.0.5H₂ O: C, 63.49; H, 7.57. Found: C, 63.54;H, 7.43.

Preparation 114β,15-Bis-(trans-2'-butenoyloxy)-3α-hydroxy-12,13-epoxytrichothec-9-ene

Repetition of the above experiment using 6 equivalents oftrans-2-butenoic acid chloride gave the title compound as a hygroscopicwhite foam. IR(KBr): 3420, 2970, 1720, 1310, 1260, 1185, 965 cm⁻¹. Inaddition, there was obtained4β(trans-2'-butenoyloxy)-3α,15-dihydroxy-12,13-epoxytrichothec-9-ene asa cream solid of m.p. 60°-62° C. IR(KBr): 3460, 2960, 1710, 1315, 1190,1105, 1080, 955 cm⁻¹.

Anal. Calcd for C₁₉ H₂₆ O₆.: C, 64.30; H, 7.53. Found: C, 64.19; H,8.06.

Preparation 1215-(2'-Methylpropenoyloxy)-3α,4β-dihydroxy-12,13-epoxytrichothec-9-ene

To a solution of 366 mg (1 mmol) of3α-O-(2'-tetrahydropyranyl)-4β,15-dihydroxy-12,13-epoxytrichothec-9-eneand 395 mg (5 mmol) of dry pyridine in 50 ml of methylene chloride(dried over 4A molecular sieves) was added with stirring 261 mg (2.5mmol) of freshly distilled 2-methylpropenoic acid chloride. The solutionwas stored at 22° C. for 17 h and was then treated with an additional261 mg (2.5 mmol) of the acid chloride. After a further 22 h at 22° C.,the solution was worked up as described in Preparation 10 and theresidue was chromatographed on 20 g of silica (Mallinckrodt SILICARCC-7). 2-Methylpropenoic acid anhydride was eluted using 1% methanol inCH₂ Cl₂. The solvent was changed to methanol to elute 230 mg of whitefoam which was hydrolyzed as described above (Preparation 10) to give189 mg of a foam. This was chromatographed on 20 g silica (MallinckrodtSILICAR CC-7) using 1% methanol in CH₂ Cl₂ as the solvent. Minorproducts were eluted and the solvent was changed to 20% methanol in CH₂Cl₂ to afford 116 mg (33%) of the title compound as a foam whichcrystallized from CH₂ Cl₂ -SKELLYSOLVE B as a pale pink solid of m.p.79°-81° C. IR(KBr): 3440, 2960, 1715, 1165, 1080, 955 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₆ O₆.0.5H₂ O: C, 63.49; H, 7.57. Found: C, 63.36;H, 7.40.

Preparation 134β,15-Bis-(2'-methylpropenoyloxy)-3α-hydroxy-12,13-epoxytrichothec-9-ene

A solution containing 3.66 g (0.01 mol) of3α-O-(2'-tetrahydropyranyl)-4β,15-dihydroxy-12,13-epoxytrichothec-9-ene,3.95 (0.05 mol) of pyridine and 2.61 g (0.025 mol) of freshly distilled2-methylpropenoic acid chloride in 250 ml of dry methylene chloride wasstirred for 16 h at 22° C. An additional 2.61 g (0.025 mol) of the acidchloride was added and stirring was continued for 6 h. The solution wasdiluted with CH₂ Cl₂ and was washed in succession with saturated aqueousNaHCO₃, brine, 1% aqueous HCl and brine. The organic phase was driedover Na₂ SO₄ and the solvent evaporated under reduced pressure to give5.36 g of an oil. This was chromatographed on 100 g of silica gel(Mallinckrodt SILICAR CC-7) using 1% methanol in CH₂ Cl₂ as the solvent.Methacrylyl anhydride was first eluted, followed by 615 mg of a foamwhich was hydrolysed as before (Preparation 10) in 67.5 ml of 95%ethanol and 13.5 ml of 1 N HCl. The usual work-up gave 590 mg of gumfrom which, by chromatography, 198 mg of the title compound was isolatedas a hygroscopic foam IR(KBr): 3500, 2960, 1720, 1165, 1080, 960 cm⁻¹ ;which was identified by its NMR spectrum. The next fraction from thischromatography afforded4β-(2'-methylpropenoyloxy)-3α,15-dihydroxy-12,13-epoxytrichothec-9-eneas colorless crystals of m.p. 175°-176° C. IR(KBr): 3510, 3460, 2500,1690, 1330, 1300, 1170, 1080, 1060, 910, 900 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₆ O₆.0.25H₂ O: C, 64.30; H, 7.53. Found: C,64.24; H, 7.14.

From the chromatographic separation of the tetrahydropyranyl ethers(above) there was next obtained 810 mg of a foam which wasre-chromatographed on fresh silica gel (20 g) using the same solventsystem to provide3α-O-[2'-tetrahydropyranyl]-15-(2'-methylpropenoyloxy)-4β-hydroxy-12,13-epoxytrichothec-9-eneas a foam.

Preparation 144β-(2'-Chloroacetoxy)-15-(2'-methylpropenoyloxy)-3α-hydroxy-12,13-epoxytrichothec-9-ene

To a stirred solution of 164 mg (0.38 mmol) of3α-O-(2'-tetrahydropyranyl)-15-(2'-methylpropenoyloxy)-4β-hydroxy-12,13-epoxytrichothec-9-ene(Preparation 13) in 25 ml of dry CH₂ Cl₂ were added in succession 36 mg(0.46 mmol) of pyridine and 78 mg (0.46 mmol) of chloroacetic anhydride.The solution was stored for 17 h at 22° C. The solution was worked up asbefore and hydrolysed as usual with 27 ml of 95% ethanol and 5.4 ml of 1N HCl. After work-up as before there was obtained a gum which wastriturated with SKELLYSOLVE B to provide a hydroscopic solid of m.p.58°-60° C. IR(KBr): 2960, 1755, 1715, 1320, 1295, 1165, 1085, 955 cm⁻¹.

Anal. Calc'd for C₂₁ H₂₇ ClO₇ : C, 59.08; H, 6.38. Found: C, 60.48; H,6.66.

Preparation 15

Following the general procedure of Preparation 5 with the bromoacetylbromide used therein replaced by an equimolar weight of the appropriateacylating agent, the following compounds were prepared:

    ______________________________________                                         ##STR19##                                                                    R.sup.1          R.sup.2                                                      ______________________________________                                        CH.sub.2 CH.sub.3                                                                             CH.sub.2 CH.sub.3                                             CH.sub.2 CH.sub.2 CH.sub.3                                                                    CH.sub.2 CH.sub.2 CH.sub.3                                    CH.sub.2 CH.sub.2 CH.sub.2 CH.sub.3                                                           CH.sub.2 CH.sub.2 CH.sub.2 CH.sub.3                            ##STR20##                                                                                     ##STR21##                                                    ______________________________________                                    

Preparation 16

If the general procedure of Preparations 6-7 is repeated with thechloroacetic anhydride used therein replaced by an equimolar amount ofthe appropriate acylating agent, the following esters may be obtained

    ______________________________________                                                 Product                                                                        ##STR22##                                                           Acylating Agent                                                                          R.sup.1        R.sup.2                                             ______________________________________                                        trifluoroacetic                                                                          CF.sub.3       CF.sub.3                                            anhydride                                                                     isobutyl chloride                                                                        CH(CH.sub.3).sub.2                                                                           CH(CH.sub.3).sub.2                                  valeryl chloride                                                                         (CH.sub.2).sub.3 CH.sub.3                                                                    (CH.sub.2).sub.3 CH.sub.3                           m-toluoyl  chloride                                                                       ##STR23##                                                                                    ##STR24##                                          p-anisyl chloride                                                                         ##STR25##                                                                                    ##STR26##                                          p-chlorobenzoyl  chloride                                                                 ##STR27##                                                                                    ##STR28##                                          phenylacetyl  chloride                                                                    ##STR29##                                                                                    ##STR30##                                          ______________________________________                                    

Preparation 17

If the general procedure of Preparation 8 is repeated with thechloroacetic anhydride used therein replaced with an equimolar amount ofthe acylating agents listed in Preparation 16, the following mixedesters may be obtained.

    ______________________________________                                         ##STR31##                                                                    R.sup.1        R.sup.2                                                        ______________________________________                                        CH.sub.3       CF.sub.3                                                       CH.sub.3       CH(CH.sub.3).sub.2                                             CH.sub.3       (CH.sub.2).sub.3 CH.sub.3                                      CH.sub.3                                                                                      ##STR32##                                                     CH.sub.3                                                                                      ##STR33##                                                     CH.sub.3                                                                                      ##STR34##                                                     CH.sub.3                                                                                      ##STR35##                                                     ______________________________________                                    

Preparation 18

Esters of the type ##STR36## where R¹ ≠R² may be prepared by a proceduresimilar to that used for Preparation 14. By using less than twoequivalents of an acylating agent listed in Preparation 16, a mixture ofmonoacylated products of the formulae ##STR37## are produced. Theseproducts may be separated chromatographically and then treated with asecond acylating agent selected from the list provided in Preparation 16(the second reagent being different than the first) to give productssuch as shown below.

    ______________________________________                                         ##STR38##                                                                    R.sup.1          R.sup.2                                                      ______________________________________                                        CF.sub.3                                                                                        ##STR39##                                                   CF.sub.3         CH(CH.sub.3).sub.2                                            ##STR40##       (CH.sub.2).sub.3 CH.sub.3                                     ##STR41##                                                                                      ##STR42##                                                   CH(CH.sub.3).sub.2                                                                              ##STR43##                                                    ##STR44##       CF.sub.3                                                     ______________________________________                                    

Preparation 19

Following the general procedures illustrated above, the following estersmay be prepared.

    ______________________________________                                         ##STR45##                                                                    R.sup.1            R.sup.2                                                    ______________________________________                                        CH.sub.3                                                                                          ##STR46##                                                  ##STR47##                                                                                        ##STR48##                                                  ##STR49##         CH.sub.2 Cl                                                 ##STR50##                                                                                        ##STR51##                                                 CH.sub.2 Cl                                                                                       ##STR52##                                                 CH.sub.2 CH.sub.2 CHClCH.sub.3                                                                   CH.sub.2 CH.sub.2 CHClCH.sub.3                             CH.sub.2 CHClCH.sub.3                                                                            CH.sub.2 CHClCH.sub.3                                       ##STR53##                                                                                        ##STR54##                                                 CCl.sub.3          CCl.sub.3                                                  CF.sub.3                                                                                          ##STR55##                                                 CH.sub.3                                                                                          ##STR56##                                                 (CH.sub.3).sub.2 CH                                                                              CH.sub.2 Cl                                                 ##STR57##         CH.sub.2 CH.sub.2 CH.sub.3                                  ##STR58##                                                                                        ##STR59##                                                  ##STR60##                                                                                        ##STR61##                                                  ##STR62##                                                                                        ##STR63##                                                  ##STR64##                                                                                        ##STR65##                                                  ##STR66##                                                                                        ##STR67##                                                 CHCH.sub.2         CHCH.sub. 2                                                 ##STR68##                                                                                        ##STR69##                                                 CCCH.sub.3         CCCH.sub.3                                                 C CH               CCH                                                         ##STR70##                                                                                        ##STR71##                                                  ##STR72##                                                                                        ##STR73##                                                  ##STR74##                                                                                        ##STR75##                                                  ##STR76##                                                                                        ##STR77##                                                  ##STR78##         CH.sub.2 Cl                                                ______________________________________                                    

EXAMPLE 1 4β,15-Diacetoxy-3α,8β-dihydroxy-12,13-epoxytrichothec-9-ene##STR79##

A mixture of 4β,15-diacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene (366mg, 1.0 mmol) of selenium dioxide (122 mg, 1.1 mmol) in 25 ml of dioxanecontaining 1 ml of water was heated to reflux for 24 h. The resultingsolution was filtered through CELITE (diatomaceous earth) and theresidue was washed with a small amount of CH₂ Cl₂. The combined filtratewas diluted with 25 ml of CH₂ Cl₂ and washed with brine. Drying over Na₂SO₄ and removal of the solvent gave 800 mg of an oil. Chromatography onsilica gel (elution with 2% methanol-CH₂ Cl₂) followed bycrystallization from CH₂ Cl₂ and diethyl ether gave 148 mg (39%) oftitle product as slightly pink crystals. Recrystallization from CH₂ Cl₂-diethyl ether gave an analytical sample: m.p. 114°-116°; IR (KBr):3435, 2965, 2943, 2905, 1730, 1715 (sh), 1365, 1248, 1080, 1061, 1035,958 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₆ O₈ : C, 59.67; H, 6.85. Found: C, 59.23; H,6.81.

EXAMPLE 215-Acetoxy-4β-chloroacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-en-8-one##STR80##

A mixture of15-acetoxy-4β-chloroacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene (0.71mmol), pyridinium chlorochromate (2.23 mg, 1.03 mmol), and anhydroussodium acetate (29 mg, 0.35 mmol) in 15 ml of CH₂ Cl₂ is stirred at roomtemperature for 2.5 h. After filtration through CELITE, the solvent isremoved under reduced pressure to give an oil. Chromatography on silicagel (elution with 0.75% methanol-CH₂ Cl₂) gives the title product.

EXAMPLE 3 4β,15-Diacetoxy-12,13-epoxytrichothec-9-en-3,8-dione ##STR81##

To a solution of 78 mg (1.0 mmol) of dimethyl sulfoxide in 2 ml of CH₂Cl₂ was added at -78° C. a 10% CH₂ Cl₂ solution of trifluoroaceticanhydride (0.6 mmol). After 10 min of stirring at -78° C. a solution of4β,15-diacetoxy-3α,8β-dihydroxy-12,13-epoxytrichothec-9-ene (76 mg, 0.2mmol) in CH₂ Cl₂ (2 ml) was added dropwise. Stirring was continued at-78° C. for 30 min and then triethylamine (101 mg, 1 mmol) was added.After an additional 10 min of stirring at -78° C., the reaction mixturewas warmed to room temperature. It was diluted with CH₂ Cl₂ (25 ml) waswashed with brine. Drying over Na₂ SO₄ and removal of the solvent gave58 mg (76%) of crystalline solid. Recrystallization from CH₂ Cl₂ anddiethyl ether gave an analytical sample of title product; m.p. 198°-199°C.; IR (KBr): 2990, 2950, 2933, 1780, 1742, 1676, 1392, 1370, 1231,1220, 1065, 1035 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₂ O₈ : C, 60.31; H, 5.86. Found: C, 60.19; H,5.78.

EXAMPLE 415-Acetoxy-4β-chloroacetoxy-3α,8β-dihydroxy-12,13-epoxytrichothec-9-ene##STR82##

A mixture of15-acetoxy-4β-chloroacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene (653mg, 1.59 mmol) and selenium dioxide (183 mg, 1.65 mmol) in 25 ml ofdioxane containing 1 ml of water was heated to reflux for 12 h. Theresulting mixture was filtered through CELITE and the filtrate wasevaporated. The residue was dissolved in CH₂ Cl₂ and washed with brine.Drying over Na₂ SO₄ and removal of the solvent gave 778 mg of a yellowoil. Chromatography on silica gel (elution with 2% methanol-CH₂ Cl₂),followed by crystallization from diethyl ether gave 275 mg (42%) ofslightly yellow crystals. m.p. 199.5°-200.5° C.; IR (KBr): 3505, 2980,2950, 1755 (sh), 1740, 1730, 1255, 1238, 1165, 1055 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₅ ClO₈ : C, 54.74; H, 6.04. Found: C, 54.30; H,5.84.

EXAMPLE 515-Acetoxy-4β-chloroacetoxy-12,13-epoxytrichothec-9-en-3,8-dione##STR83##

To a solution of dimethyl sulfoxide (0.192 ml, 2.71 mmol) in 3 ml of CH₂Cl₂ was added at -78° C. neat trifluoroacetic anhydride (0.217 ml, 1.63mmol). After 10 min at -78° C., a CH₂ Cl₂ (5 ml) solution of15-acetoxy-4β-chloroacetoxy-3α,8β-dihydroxy-12,13-epoxytrichothec-9-ene(245 mg, 0.588 mmol) was added dropwise. The mixture was stirred at -78°for 45 min, followed by a dropwise addition of triethylamine (0.381 ml,2.71 mmol). The reaction mixture was kept at -78° C. for 10 min and thenallowed to warm to room temperature. The resulting solution was dilutedwith CH₂ Cl₂ (15 ml) and washed with brine. Drying over Na₂ SO₄ andremoval of the solvent gave 303 mg of oil. Crystallization from diethylether yielded 160 mg (66 %) of yellow crystals; m.p. 169.5°-171° C. IR(KBr): 3000, 2962, 1772, 1747, 1678, 1232, 1219, 1156, 1052 cm⁻¹.

Anal. Calc'd for C₁₉ H₂₁ ClO₈ : C, 55.28; H, 5.12. Found: C, 54.31; H,5.06.

EXAMPLE 6

If the general procedure of Examples 1 and 4 are repeated with the4β,15-diacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene or15-acetoxy-4β-chloroacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene usedtherein replaced by an equimolar amount of a 3α-hydroxy ester listedbelow, there is produced the corresponding 3,8-dihydroxy ester product.

    ______________________________________                                         ##STR84##                                                                     ##STR85##                                                                    R.sup.1            R.sup.2                                                    ______________________________________                                        CH.sub.3                                                                                          ##STR86##                                                  ##STR87##                                                                                        ##STR88##                                                  ##STR89##         CH.sub.2 Cl                                                 ##STR90##                                                                                        ##STR91##                                                 CH.sub.2 Br        CH.sub.2 Br                                                CH.sub.2 Cl        CH.sub.2 Cl                                                CH.sub.2 Cl                                                                                       ##STR92##                                                 CH.sub.2 CH.sub.3  CH.sub.2 CH.sub.3                                          CH.sub.2 CH.sub.2 CH.sub.3                                                                       CH.sub.2 CH.sub.2 CH.sub.3                                 CH.sub.2 CH.sub.2 CH.sub.2 CH.sub.3                                                              CH.sub.2 CH.sub.2 CH.sub.2 CH.sub.3                        CH.sub.2 CH.sub.2 CHClCH.sub.3                                                                   CH.sub.2 CH.sub.2 CHClCH.sub.3                             CH.sub.2 CHClCH.sub.3                                                                            CH.sub.2 CHClCH.sub.3                                       ##STR93##                                                                                        ##STR94##                                                 CCl.sub.3          CCl.sub.3                                                  CF.sub.3           CF.sub.3                                                    ##STR95##                                                                                        ##STR96##                                                 (CH.sub.3).sub.2 CH                                                                              (CH.sub.3).sub.2 CH                                         ##STR97##                                                                                        ##STR98##                                                  ##STR99##                                                                                        ##STR100##                                                 ##STR101##                                                                                       ##STR102##                                                CF.sub.3                                                                                          ##STR103##                                                CH.sub.3                                                                                          ##STR104##                                                (CH.sub.3).sub.2 CH                                                                              CH.sub.2 Cl                                                 ##STR105##        CH.sub.2 CH.sub.2 CH.sub.3                                  ##STR106##                                                                                       ##STR107##                                                 ##STR108##                                                                                       ##STR109##                                                 ##STR110##                                                                                       ##STR111##                                                 ##STR112##                                                                                       ##STR113##                                                 ##STR114##                                                                                       ##STR115##                                                 ##STR116##                                                                                       ##STR117##                                                 ##STR118##                                                                                       ##STR119##                                                 ##STR120##                                                                                       ##STR121##                                                 ##STR122##        CH.sub.2 Cl                                            

EXAMPLE 7

If the general procedure of Example 2 is repeated with the15-acetoxy-4β-chloroacetoxy-3α-hydroxy-12,13-epoxytrichothec-9-ene usedtherein replaced by an equimolar amount of a 3,8-dihydroxy ester listedbelow, there is produced the corresponding 3α-hydroxy-8-keto product.

    ______________________________________                                         ##STR123##                                                                    ##STR124##                                                                   R.sup.1            R.sup.2                                                    ______________________________________                                        CH.sub.3                                                                                          ##STR125##                                                 ##STR126##                                                                                       ##STR127##                                                 ##STR128##        CH.sub.2 Cl                                                 ##STR129##                                                                                       ##STR130##                                                CH.sub. 2 Br       CH.sub.2 Br                                                CH.sub.2 Cl        CH.sub.2 Cl                                                CH.sub.2 Cl                                                                                       ##STR131##                                                CH.sub.2 CH.sub.3  CH.sub.2 CH.sub.3                                          CH.sub.2 CH.sub.2 CH.sub.3                                                                       CH.sub.2 CH.sub.2 CH.sub.3                                 CH.sub.2 CH.sub.2 CH.sub.2 CH.sub.3                                                              CH.sub.2 CH.sub.2 CH.sub.2 CH.sub.3                        CH.sub.2 CH.sub.2 CHClCH.sub.3                                                                   CH.sub.2 CH.sub.2 CHClCH.sub.3                             CH.sub.2 CHClCH.sub.3                                                                            CH.sub.2 CHClCH.sub.3                                       ##STR132##                                                                                       ##STR133##                                                CCl.sub.3          CCl.sub.3                                                  CF.sub.3           CF.sub.3                                                    ##STR134##                                                                                       ##STR135##                                                (CH.sub.3).sub.2 CH                                                                              (CH.sub.3).sub.2 CH                                         ##STR136##                                                                                       ##STR137##                                                 ##STR138##                                                                                       ##STR139##                                                 ##STR140##                                                                                       ##STR141##                                                CF.sub.3                                                                                          ##STR142##                                                CH.sub.3                                                                                          ##STR143##                                                (CH.sub.3).sub.2 CH                                                                              CH.sub.2 Cl                                                 ##STR144##        CH.sub.2 CH.sub.2 CH.sub.3                                  ##STR145##                                                                                       ##STR146##                                                 ##STR147##                                                                                       ##STR148##                                                 ##STR149##                                                                                       ##STR150##                                                 ##STR151##                                                                                       ##STR152##                                                 ##STR153##                                                                                       ##STR154##                                                 ##STR155##                                                                                       ##STR156##                                                 ##STR157##                                                                                       ##STR158##                                                 ##STR159##                                                                                       ##STR160##                                                 ##STR161##        CH.sub.2 Cl                                                ______________________________________                                    

EXAMPLE 8

If the general procedures of Examples 3 and 5 are repeated with the4β,15-diacetoxy-3α,8β-dihydroxy-12,13-epoxytrichothec-9-ene or15-acetoxy-4β-chloroacetoxy-3α,8β-dihydroxy-12,13-epoxytrichothec-9-eneused therein replaced by an equimolar amount of a 3,8-dihydroxy esterlisted below, there is produced the corresponding 3,8-diketo product.##STR162##

This invention is capable of industrial application.

We claim:
 1. A compound of the formula ##STR163## wherein R¹ and R² areeach independently (lower)alkyl; halo(lower)alkyl; alkenyl of theformula --CR³ ═CR⁴ R⁵ in which R³ is hydrogen, (lower)alkyl or1'-halo(lower)alkyl and R⁴ and R⁵ are each independently hydrogen or(lower)alkyl; alkynyl of the formula --C.tbd.CR⁶ in which R⁶ is hydrogenor (lower)alkyl; or a radical of the formula

    Ar--(CH.sub.2).sub.m --

in which m is 0 or an integer from one to four and Ar is ##STR164##wherein R⁷, R⁸ and R⁹ are each independently hydrogen, halogen,(lower)alkyl or (lower)alkoxy.
 2. A compound as claimed in claim 1wherein R¹ and R² are each independently (lower)alkyl, halo(lower)alkylor --CR³ ═CR⁴ R⁵ in which R³, R⁴ and R⁵ are each independently hydrogenor (lower)alkyl.
 3. A compound as claimed in claim 1 wherein R¹ and R²are each independently (lower)alkyl, --CH₂ Cl or ##STR165##
 4. Thecompound of claim 1 wherein R¹ and R² are each methyl.
 5. The compoundof claim 1 wherein R¹ is --CH₃ and R² is --CH₂ Cl.
 6. The compound ofclaim 1 wherein R¹ is --CH₃ and R² is ##STR166##
 7. The compound ofclaim 1 wherein R¹ and R² are each ##STR167##